Project Summary/Abstract Obstructive sleep apnea (OSA) is a serious health issue, characterized by repeated episodes (100s/night) of airway obstruction and apnea (no airflow), and increased risk of cardiovascular disease. Normally, activity of the tongue (particularly genioglossus, primary tongue protruder) stiffens the airway to keep it open. Notably, the tongue receives inspiratory drive derived from the preBtzinger Complex (preBtC; generates the drive) and is transmitted via inspiratory XII premotoneurons (hypoglossal preMNs) to XII MNs that innervate the tongue. During sleep, reduced XII MN activity is hypothesized to result from activation of sleep-specific cholinergic modulation at XII MNs, which has then been implicated in airway collapse in OSA. This project will explore the hypothesis that cholinergic drive activates inhibitory G-protein coupled inward rectifying potassium (GIRK) receptors, leading to reduced XII MN activity by acting directly on XII MNs, or by reducing excitability and output of XII inspiratory preMNs and thus indirectly acting at XII MNs. This project uses in vitro electrophysiology, in combination with the PI?s recent discovery that XII inspiratory preMNs originate from a neuronal class derived from the transcription factor Dbx1, to address the following two specific aims: Aim 1) Elucidate cholinergic mechanisms of action at XII MNs across postnatal maturation. Using rhythmic medullary slice preparations from 0-14 days postnatal (P0-14) and up to adult mice, cholinergic mechanisms of action, such as via GIRK channels, will be explored at XII MNs by recording the whole XII nerve output, and at single XII MNs using whole cell electrophysiology techniques. Aim 2) Elucidate cholinergic mechanisms of action at Dbx1 XII inspiratory preMNs across postnatal maturation. Using rhythmic medullary slice preparations from P0-14 and up to adult mice, cholinergic mechanisms of action will be elucidated at individual Dbx1 XII inspiratory preMNs using whole cell electrophysiology techniques. These studies will contribute excellent training opportunities for graduate and professional students, thus providing scientific training to the next generation of clinicians. Furthermore, the expected outcomes will advance understanding of cellular and synaptic cholinergic mechanisms that are key determinants of XII MN inspiratory activity. This basic information will be essential to develop strategies to explore in vivo the cholinergic mechanisms that contribute to state-dependent reductions in airway tone. These data will contribute to the mission of the National Heart Lung, and Blood Institute of improving the health and quality of Americans suffering from sleep disordered breathing.